Major hypothesis of this project: The endogenous opioid, cannabinoid, and CCK systems mutually interact via hippocampal interneurons and thereby jointly regulate the excitability of pyramidal cells. Low frequency neuronal population oscillations (especially theta ?, 4-12 Hz, and gamma ?, 30-80 Hz, rhythms) represent brain states that are crucial for high order cognitive processing. Activity in GABAergic basket cell microcircuits is required for several forms of rhythm generation, yet many questions regarding the underlying cellular mechanisms are unresolved, largely because available tools are inadequate for selectively investigating the sparse and widely dispersed cellular circuits. Model phenomena, called low frequency oscillations (LFOs), capture some of the properties of the much more complex in vivo brain rhythms. The present application will use new optogenetic methods to address questions about mechanisms of LFOs at the microcircuit level. The goal is to elucidate fundamental cell circuit properties that may help illuminate the in vivo phenomena. Molecular biological tools will be used to introduce light-sensitive molecules (opsins) into targeted cell groups via viral vectors. The vectors contain a double-floxed, inverted gene for a fusion protein consisting of an opsin plus a fluorescent marker protein. Vectors are introduced into the brains of transgenic mice expressing Cre-recombinase under the control of cell-specific promoters. We will target the opsins to either the parvalbumin (PV) - expressing, or cholescystokinin (CCK) - expressing interneurons in hippocampus, or acetylcholine (ACh) - expressing cells in the medial septum. Depending on its molecular properties, light-activation of a particular opsin will either excite or inhibit the cell expressing it. We will use the excitatory opsin, Channelrhodopsin2, and the inhibitory opsin, Halorhodopsin in the proposed experiments. Flashes of light of an appropriate wavelength will either excite or inhibit defined networks of cells, even though the cells are scattered in the tissue. Optogenetic methods will complement high resolution electrophysiological analysis of individual neurons. In the hippocampus, the mu-opioid receptor, ?OR, or the cannabinoid receptor, CB1R, are segregated at high density on the PV or CCK cells, respectively. We will ask how the individual microcircuits regulated by ?ORs and CB1Rs interact to foster inhibitory LFOs. The Specific Aims are to: Aim #1: Test the hypothesis that CCK and PV cells mutually influence each other. Aim #2: Test the hypothesis that CCK and PV cell IPSPs collectively generate LFO-LFPs. Aim #3: Test the hypothesis that LFO-IPSCs and LFO-LFPs can be triggered by endogenous ACh. PUBLIC HEALTH RELEVANCE: Health Relevance: Disordered function of inhibitory microcircuits has been implicated in psychiatric disorders such as schizophrenia, autism, and Alzheimer's Disease, among others. In addition, opiate and cannabinoid drugs have actions that interact in affecting behavior, and yet the ways in which they interact is not known. A detailed understanding of the cross-talk between PV and CCK cell microcircuits will impact on important neurological problems.